Project Summary Coccidioidomycosis affects residents in the southwestern United States, northern Mexico and scattered areas of South America. An estimated 150,000 people in the United States become infected with Coccidioides annually. There is an urgent unmet need to develop better chemotherapies and a vaccine against Coccidioides infection. This proposed study is to optimize protective efficacy and delineate immune mechanisms of a newly developed multivalent vaccine against coccidioidomycosis. The vaccine consists of a recombinant chimeric polypeptide antigen (rCPA2) composed of the most immunogenic fragments of 4 previously identified Coccidioides antigens and 5 predicted T-cell epitopes. Preliminary studies demonstrate robust and long-lasting adaptive T-cell responses following vaccination of human MHC II-expressing HLA-DR4 transgenic mice with glucan-chitin particles (GCPs) ?loaded? with rCPA2. Furthermore, optimized GCP-rCPA2 vaccine affords the best protective efficacy among 5 tested adjuvant formations and comparable to a live, attenuated ?T vaccine. The rCPA+GCP vaccine is well processed by macrophages and dendritic cells at the vaccination sites and mediates differential immune gene expressions leading to enhanced antigen presentation and Th1- and Th17- mixed polarization. Th17 immunity has shown to be indispensable for vaccine immunity against coccidioidomycosis. The central hypothesis of this proposal is that an optimized rCPA2 antigen loaded in GCP adjuvant/delivery system with an FDA-approved carrier can elicit a broad spectrum of protective immunity for humanized mice against infection with both species of Coccidioides. The goal is to investigate the protective mechanisms of the optimized GCP-rCPA2 vaccine using humanized murine models of coccidioidomycosis and human immune cells. There are 3 Specific Aims: Aim 1 is to create an optimized rCPA2 that can induce a broad spectrum and durable protective immunity against both species of Coccidioides. Human T-cell epitopes on rCPA2 will be mapped for 3 common human MHC II alleles. There is considerable morphological and genomic diversity among different isolates of C. posadasii and C. immitis. The optimized antigen will include dominant epitopes derived from both species of Coccidioides for multiple HLA alleles. Protective efficacy of the optimized vaccine will be tested to against two selected isolates of each Coccidioides species that present with the least homology of the antigens. Aim 2 is to optimize the GCP adjuvant formulation for eliciting a protective Th1- and Th17-mixed response against Coccidioides infection using both human and mouse APCs and CD4+ T cells. An optimized GCP adjuvant/delivery system will be created to maximize protective efficacy. Aim 3 To study the vaccine induced protective mechanism that guides Th1 and Th17 responses using murine models of coccidioidomycosis. Upon completion of this project an optimized and protective GCP2-rCPA2 vaccine will be identified for advancement to clinical trials for assessment of it safety and protective efficacy against coccidioidomycosis.